Histone modifications are thought to regulate gene expression by altering the accessibility of DNA to the transcription machinery. They are implicated in cell reprogramming, development, and disease, but it remains unclear if they actively control gene expression, or simply go along for the ride. Our goal is to help answer this important question by using technology we are developing to directly image histone modification dynamics in single living cells and at single copy genes with record-breaking spatio-temporal resolution. My lab at Colorado State University is well-prepared to use our new technology to tackle tough biological questions. In particular, we have collected a substantial amount of preliminary data that demonstrates the feasibility of our strategy. By combining the best of FabLEM (Fab based Live Endogenous Modification labeling) and single molecule tracking, we have made a number of key innovations that together lay the foundation for multiplexed imaging of histone modifications with super-resolved spatial resolution (<30 nm) and high-throughput. Key questions we plan on addressing include: (1) How do histone modifications dynamically regulate the transcriptional output of single-copy genes? (2) What is the timescale of histone modification turnover and does this impact cell inheritance? and (3) How do histone modifications interact with specific oncogenes and their targets? A theme that unifies our approach to all of these questions is the pre-marking of genes together with high-resolution imaging of specific histone modifications. This ability gives us a unique advantage and allows us to directly attack our questions with direct imaging not possible before.
Histone modifications correlate with gene activity, but whether or not they causally regulate genes remains unclear due to the poor spatio-temporal resolution of available techniques for quantifying modification dynamics in vivo. We will develop and apply technology to image histone modifications in living cells with unprecedented spatio-temporal resolution to help resolve key questions of histone modification causality in transcriptional regulation, epigenetic inheritance, and cancer progression.
| Lyon, Kenneth; Stasevich, Timothy J (2017) Imaging Translational and Post-Translational Gene Regulatory Dynamics in Living Cells with Antibody-Based Probes. Trends Genet 33:322-335 |
| Ecklund, Kari H; Morisaki, Tatsuya; Lammers, Lindsay G et al. (2017) She1 affects dynein through direct interactions with the microtubule and the dynein microtubule-binding domain. Nat Commun 8:2151 |
